The objectives of the proposed research are to obtain a detailed understanding of the physiological functioning and metabolic regulation of a highly branched biosynthetic pathway and to learn the physiochemical mechanisms of catalysis and regulation of this system. The enzyme system chosen is the phosphoribosylpyrophosphate synthetase of Salmonella typhimurium, which catalyzes an ususual pyrophosphate group transfer reaction and which is required for synthesis of a large group of nucleic acid precursors, two amino acids, and two coenzymes. Regulation of synthesis and activity of the enzyme will be investigated by study of appropriate mutant strains and by kinetic and physical studies of the highly purified enzyme. Evaluation of the physiological functioning of PRPP synthetase in living cells has been achieved by determination of the role of ribose-5-P in regulating PRPP levels. Isolation of mutants with defects in PRPP synthetase activity is currently under way. Characterization of these and similar mutants should clarify the control of synthesis and metabolic functions of PRPP synthetase in vivo. The mechanism of catalysis will be investigated by a variety of kinetic, chemical modification, and magnetic resonance techniques. The results will relate to the mechanism of catalysis by many nucleotide-dependent enzymes. In the broadest sense this research is directed toward a better understanding of the regulation of cellular metabolism, the mechanism of biochemically important transfer reactions, and biochemical mechanisms of metabolic diseases, such as hyperuricemia, which are characterized by derangements in nucleotide metabolism and which involve PRPP synthetase in some cases.